Heart disease is one of the most serious health concerns in the western world. It is estimated that 61 million Americans (nearly 1 in 5 men and women) have one or more types of cardiovascular disease (National Health and Nutrition Examination Survey III, 1988-1994, Center of Disease Control and the American Heart Association). Widespread conditions include coronary heart disease (12.4 million), congenital cardiovascular defects (1 million), and congestive heart failure (4.7 million). A central challenge for research in regenerative medicine is to identify and develop drugs that can help reconstitute cardiac function in these conditions.
The development of new drugs is hampered by the lack of suitable cell-based in vitro systems that resemble a diseased tissue, for example myopathic cardiac cells. A variety of attempts to obtain immortalized cardiac myocytes are described in Sen et al., J. Biol. Chem. 263 (1988), 19132-19136; Gartside and Hauschka in “The Development and Regenerative Potential of Cardiac Muscle”, eds. Oberpriller et al., Harwood, N.Y., 1991, 7941-7948; Jaffredo et al., Exp. Cell. Res. 192 (1991), 481-491; Wang et al., In Vitro Cell Dev. Biol. 27 (1991), 63-74; Katz et al., Am. J. Physiol. 262 (1992), 1867-1876); Engelmann et al., J. Mol. Cell Cardiol. 25 (1993), 197-213; Borisov and Claycomb, Ann. NY Acad. Sci. 752 (1995), 80-91; Jahn et al., J. Cell Sci. 109 (1996), 397-407. However, the cardiac phenotype of the cells so obtained either is not stable, or the cells loose their ability to proliferate. Furthermore, a cell line of murine immortalized, atrial cardiomyocytes has been described, which maintains features of differentiation and capability of proliferation over a longer period of time (Claycomb et al., Proc. Natl. Acad. Sci. USA 95 (1998), 2979-2984).
As an in vitro heart disease model system based on non-transformed cardiomyocytes often preparations of heart cells isolated from mouse or rat are used; see Chlopcikova et al., Biomed. Pap. Med. Fac. Univ. Palacky Olomouc Czech. Repub. 145 (2001), 49-55. These cells maintain their differentiated phenotype for a few days only. Furthermore, these primary cultures are not homogeneous but contain different cell types and vary for each preparation. A particular problem consists in the contamination of the cardiomyocytes by other cell types present in the heart, in particular fibroblasts, which in contrast to resting cardiomyocytes strongly proliferate and cannot entirely be eliminated from the culture. Some receptors expressed by cardiomyocytes as well as by non-cardiomyocyte cells and cardiomyocytes secrete molecules which interact with receptors from non-cardiomyocytes, as well as vice versa non-cardiomyocytes secrete molecules which bind to receptors from cardiomyocytes.
Accordingly, in view of the quite laborious preparation of cells of tissue or organs, which could serve as model system for a disease phenotype in vitro, transgenic animal models are still used such as the transgenic animal model of heart failure described in international application WO97/36477 or for human cardiomyopathy described in German patent application No. 198 15 128. Recently, a transgenic animal model to produce cardiac hypertrophy in transgenic mice has been described in U.S. Pat. No. 6,657,104.
However, these test procedures have the disadvantage that they require the use of a large number of live mammals, in particular rats and mice, and obviously are not amenable to high throughput screening.
Thus, there is a need for cell-based in vitro assay systems that can be easily performed and give reliable results. The solution to said technical problem is achieved by providing the embodiments characterized in the claims, and described further below.